Ehrlichiosis and anaplasmosis are a group of emerging zoonotic tick-borne infections caused by Ehrlichia and Anaplasma species, respectively. These obligate intracellular, gram-negative rickettsial organisms infect leukocytes and cause a potentially serious febrile illness in humans.

Human granulocytic anaplasmosis (HA) is caused by Anaplasma phagocytophilum, which is transmitted through the bite of an infected Ixodes species tick. The epidemiology of this infection in the United States is very much like that of Lyme disease (caused by Borrelia burgdorferi) and babesiosis (caused primarily by Babesia microti), which all have the same tick vector. HA is most prevalent in the upper Midwest and in other areas of the United States that are endemic for Lyme disease.

Human monocytic ehrlichiosis (HE) is caused by Ehrlichiachaffeensis, which is transmitted by the Lone Star tick, Amblyommaamericanum. Most cases of HE have been reported from the southeastern and south-central regions of the United States. E ewingii, the known cause of canine granulocytic ehrlichiosis, can occasionally cause an HE-like illness in humans. Clinical features and laboratory abnormalities are similar to those of E chaffeensis infection, and antibodies to E ewingii cross-react with current serologic assays for detection of antibodies to E chaffeensis.

Most recently, Mayo Clinic Laboratories detected a new species of Ehrlichia in patients with exposure to ticks in Wisconsin and Minnesota. This organism is most closely related to E muris and has therefore been referred to as the E muris-like agent or EMLA. The name E muris eauclairensis has recently been proposed after the city in which the first case was described. E muris eauclairensis causes a similar disease to ehrlichiosis due to E chaffeensis and E ewingii, and may cause more severe disease in immunocompromised hosts.

Most cases of anaplasmosis and ehrlichiosis are subclinical or mild, but infection can be severe and life-threatening in some individuals. Fever, fatigue, malaise, headache, and other "flu-like" symptoms, including myalgias, arthralgias, and nausea, occur most commonly. Central nervous system involvement can result in seizures and coma.

Diagnosis may be difficult since the patient's clinical course is often mild and nonspecific. This symptom complex is easily confused with other illnesses such as influenza, or other tick-borne zoonoses such as Lyme disease, babesiosis, and Rocky Mountain spotted fever. Clues to the diagnosis of ehrlichiosis in an acutely febrile patient after tick exposure include laboratory findings of leukopenia or thrombocytopenia and elevated serum aminotransferase levels. However, while these abnormal laboratory findings are frequently seen, they are not specific. Rarely, intra-granulocytic or monocytic morulae may be observed on peripheral blood smear, but this is not a reliable means of diagnosing cases of human ehrlichiosis or anaplasmosis.

Definitive diagnosis is usually accomplished through PCR and serologic methods. Serologic testing is done primarily for confirmatory purposes, by demonstrating a 4-fold rise or fall in specific antibody titers to Ehrlichia species or Anaplasma antigens. There is not currently a commercially available specific serologic test for E muris eauclairensis, but cross-reactivity with the other Ehrlichia species by serology may be detected.

PCR techniques allow direct detection of pathogen-specific DNA from patients' whole blood and is the preferred method for detection during the acute phase of illness. The Mayo PCR assay is capable of detecting and differentiating A phagocytophilum, E chaffeensis, E ewingii, and E muris eauclairensis.

It is important to note that concurrent infection with A phagocytophilum, Borrelia burgdorferi, and Babesia microti is not uncommon as these organisms share the same Ixodes tick vector, and additional testing for these pathogens may be indicated.

This test may detect DNA of Ehrlichia canis (reported to cause asymptomatic infection in Venezuela only).

This PCR test does not detect DNA of Rickettsia (formerly Ehrlichia) sennetsu, which has been reported to cause a rare mononucleosis-like illness in humans (in Japan and Malaysia).

Supportive Data

The following validation data supports the use of this assay for clinical testing.

Accuracy/Diagnostic Sensitivity and Specificity:

Results from this real-time PCR assay on the LightCycler (LC PCR) were compared to those generated using conventional PCR assay for Anaplasma phagocytophilum on 127 unique, archived whole blood specimens (26 positive and 99 negative specimens by conventional PCR). Using the conventional PCR as the gold standard, the diagnostic sensitivity and specificity for detection of Anaplasma phagocytophilum were 100%. In addition, 12 known Ehrlichia chaffeensis isolates and 2 Ehrlichia ewingii isolates (reference strains) were tested by the LC PCR and were positive.

Supplemental Data (Spiking Studies):

To supplement the above data, 30 negative whole blood samples were spiked with Anaplasma phagocytophilum positive control plasmid at the limit of detection (LoD) (10 copies/microL). The 30 spiked specimens were run in a blinded manner along with 30 negative (nonspiked) specimens. 100% of the spiked specimens were positive, and 100% of the nonspiked specimens were negative.

Analytical Sensitivity/Limit of Detection (LoD):

The lower LoD of this assay for each of the species in EDTA blood is as follows:

Fifty whole blood specimens from normal donors were tested and found to be negative for targeted or detectable Ehrlichia and Anaplasma species.

Reportable Range:

This is a qualitative assay, and results are reported as either negative or positive for targeted Ehrlichia/Anaplasma species (positive for Anaplasma phagocytophilum, Ehrlichia chaffeensis, Ehrlichia muris eauclairensis or Ehrlichia ewingii).